CRISPR-Cas systems in antiviral and antimicrobial defence: recent advances, challenges, and biodefense implications

Abstract

CRISPR–Cas systems have evolved from a bacterial immune mechanism into a versatile platform for precision medicine and molecular diagnostics. This review synthesizes design principles and failure modes for three main application areas: (i) antivirals, where conservation-aware, multiplexed guides and delivery methods (AAV, LNP, or RNA RNP) are crucial for sustained suppression; (ii) precision antimicrobials, which allow for strain-specific killing or genetic disarming (chromosomal cuts, plasmid curing, CRISPRi) with minimal impact on commensal microbes; and (iii) Cas12/Cas13 diagnostics, promoting rapid, contamination-controlled, sample-to-answer workflows. In various use cases, we focus on specificity mapping (GUIDE-seq/CIRCLE-seq and related assays), escape management (including multiplexing, ortholog switching, and Acr awareness), and governance practices (such as assay hygiene, data provenance, and biosafety). We also address delivery challenges, matrix effects, and surveillance-driven retargeting as key factors influencing success. The aim is to create a playbook that guides choices of effectors, guide design and validation, delivery routes, and quality control measures, enabling the translation of programs from research to real-world deployment.